Production of cloned goats by transfer of nuclei from transfected caprine mammary gland epithelial cells

Abstract:

In this study, we evaluated the development potential of caprine mammary gland epithelial cells (CMGECs) after transfection and nuclear transfer into enucleated, ovulated oocytes. We first isolated CMGECs from udders of lactating goats which were transfected with expression plasmid for human lacterrin and selected by G418. Then we chose sixteen neomycin resistant lines and induced them with prolactin for the expression of human lactoferrin checked by Western blotting. The donor cells, expressing human lactoferrin of 75 kD, were fused and activated with enucleated ovulated oocytes. Pronuclear-stage reconstructed embryos were transferred into the oviducts of 16 recipient goats. There were fourteen (87.5%), thirteen (81.3%), and ten (62.5%) pregnancies confirmed pregnant by ultrasound on Day 30, 60, and 90, respectively. Three recipients carried the pregnancies to term and delivered one goat each. Nested PCR-RFLP analysis confirmed that all of the kids were clones of the donor cells. These results demonstrated that CMGECs after transfection remain totipotent for nuclear transfer.